Enzymes are biocatalysts which regulate many of the biochemical reactions that occur naturally in living organisms. Enzymes are used in many fields, such as in the tanning, detergent, food and pharmaceutical industries. In the traditional method of producing enzymes, an enzyme precipitate is dissolved in water. High potency enzyme-in-water solutions (concentrated solutions having high enzyme activity), however, may be storage unstable; the enzyme precipitates out of solution and/or is heat sensitive. U.S. Pat. No. 3,242,056 discloses the use of aqueous polyol solutions to help prevent thermal instability of lysozyme. U.S. Pat. No. 4,497,897 discloses a stable liquid proteinase from Subtilisin Carlsberg containing propylene glycol, calcium ion and carboxylate salt. U.S. Pat. No. 4,519,934 discloses a stable liquid alpha-amylase from Bacillus licheniformis dispersed in propylene glycol.
A liquid enzyme product comprising alpha-amylase from Bacillus licheniformis in aqueous solution is desirable since it can be put directly to its industrial use, such as in liquid detergents, without the inconvenience of first resolubilizing a dry, enzyme precipitate in water and/or without the concern of solvent (e.g. propylene glycol) compatibility. Also, enzyme dust from a dry product is avoided. Thus, researchers have long sought methods to maintain this enzyme in aqueous solution, particularly high potency (highly concentrated) solutions. Although the prior art mentioned in the paragraph above shows the use of various solvents (e.g. propylene glycol), never is there obtained an aqueous enzyme product of high enzyme activity where this enzyme stays in solution.
For instance, alpha-amylase can be obtained from culturing various microorganisms, such as Bacillus subtilis or Bacillus licheniformis in a suitable nutrient medium. B. subtilis produces alpha-amylase that is physically stable, i.e. the enzyme will not appreciably precipitate out of an aqueous solution even when the activity is 3 million MWU/ml (Modified Wohlgemuth Units per milliliter), or higher. However, these alpha-amylase solutions are heat sensitive, and maltodextrin is added to enhance thermostability. Similarly, Klesov et al in "Substrate Thermostabilization of Soluble and Immobilized Glucoamylase", Department of Chemistry, Moscow State University, Biokhimiya, Vol. 44 (6), pages 1084-92 (1979), disclose thermostabilization of a solution of glucoamylase from Aspergillus niger with maltodextrin. On the other hand, alpha-amylase obtained from Bacillus licheniformis is heat stable (see for instance, Volesky et al, Microbial Enzymes: Production, Purification, and Isolation, Volume 2, Issue 2, page 120, 1985) but it is not especially physically stable in an aqueous solution at high concentrations.